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Impaired Vitamin D Signaling in Endothelial Cell Leads to an Enhanced Leukocyte-Endothelium Interplay: Implications for Atherosclerosis Development.

Bozic M, Álvarez Á, de Pablo C, Sanchez-Niño MD, Ortiz A, Dolcet X, Encinas M, Fernandez E, Valdivielso JM - PLoS ONE (2015)

Bottom Line: Knockdown of VDR in endothelial cells (shVDR) led to endothelial cell activation, characterized by upregulation of VCAM-1, ICAM-1 and IL-6, decreased peripheral blood mononuclear cell (PBMC) rolling velocity and increased PBMC rolling flux and adhesion to the endothelium. shVDR cells showed decreased IκBα levels and accumulation of p65 in the nucleus compared to shRNA controls.In vivo, deletion of VDR led to significantly larger aortic arch and aortic root lesions in apoE-/- mice, with higher macrophage content. apoE-/-VDR-/-mice showed higher aortic expression of VCAM-1, ICAM-1 and IL-6 when compared to apoE-/-VDR+/+ mice.Our data demonstrate that lack of VDR signaling in endothelial cells leads to a state of endothelial activation with increased leukocyte-endothelial cell interactions that may contribute to the more severe plaque accumulation observed in apoE-/-VDR-/- mice.

View Article: PubMed Central - PubMed

Affiliation: Nephrology Research Department, IRB Lleida, Lleida, Spain.

ABSTRACT
Endothelial cell activation leading to leukocyte recruitment and adhesion plays an essential role in the initiation and progression of atherosclerosis. Vitamin D has cardioprotective actions, while its deficiency is a risk factor for the progression of cardiovascular damage. Our aim was to assess the role of basal levels of vitamin D receptor (VDR) on the early leukocyte recruitment and related endothelial cell-adhesion-molecule expression, as essential prerequisites for the onset of atherosclerosis. Knockdown of VDR in endothelial cells (shVDR) led to endothelial cell activation, characterized by upregulation of VCAM-1, ICAM-1 and IL-6, decreased peripheral blood mononuclear cell (PBMC) rolling velocity and increased PBMC rolling flux and adhesion to the endothelium. shVDR cells showed decreased IκBα levels and accumulation of p65 in the nucleus compared to shRNA controls. Inhibition of NF-κB activation with super-repressor IκBα blunted all signs of endothelial cell activation caused by downregulation of VDR in endothelial cells. In vivo, deletion of VDR led to significantly larger aortic arch and aortic root lesions in apoE-/- mice, with higher macrophage content. apoE-/-VDR-/-mice showed higher aortic expression of VCAM-1, ICAM-1 and IL-6 when compared to apoE-/-VDR+/+ mice. Our data demonstrate that lack of VDR signaling in endothelial cells leads to a state of endothelial activation with increased leukocyte-endothelial cell interactions that may contribute to the more severe plaque accumulation observed in apoE-/-VDR-/- mice. The results reveal an important role for basal levels of endothelial VDR in limiting endothelial cell inflammation and atherosclerosis.

No MeSH data available.


Related in: MedlinePlus

VDR ablation affects atheroma composition in apoE-/- mice.Tissue sections of aortic arch (A, B) and aortic root (D, E) from apoE-/- and apoE-/-VDR-/- mice were stained with anti-Mac-3 to quantify plaque macrophage content. (C) Staining intensity and % of Mac-3 positive cells in the aortic arch were graded semiquantitatively and histological scores were obtained from every sample, as explained in Materials and Methods. (F) Quantification of Mac-3 staining in the aortic root is shown as percentage of stained area relative to the area occupied by atheroma. The photomicrographs (20x magnification) show representative images of aortic arch and aortic root sections. Differences between groups were evaluated by the Student t test. Data presented are mean ± SEM of 10 mice/group. *p<0.05, *#p<0.001 vs. apoE-/- mice.
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pone.0136863.g005: VDR ablation affects atheroma composition in apoE-/- mice.Tissue sections of aortic arch (A, B) and aortic root (D, E) from apoE-/- and apoE-/-VDR-/- mice were stained with anti-Mac-3 to quantify plaque macrophage content. (C) Staining intensity and % of Mac-3 positive cells in the aortic arch were graded semiquantitatively and histological scores were obtained from every sample, as explained in Materials and Methods. (F) Quantification of Mac-3 staining in the aortic root is shown as percentage of stained area relative to the area occupied by atheroma. The photomicrographs (20x magnification) show representative images of aortic arch and aortic root sections. Differences between groups were evaluated by the Student t test. Data presented are mean ± SEM of 10 mice/group. *p<0.05, *#p<0.001 vs. apoE-/- mice.

Mentions: To determine how VDR affects the nature of plaques, we further investigated atherosclerotic plaque composition. The increased lesion areas of the aortic arch and aortic root of apoE-/-VDR-/- mice correlated with increased macrophage (Mac-3) infiltration in both aortic regions (Fig 5A–5F). Immunoreactivity for monocyte chemoattractant protein (MCP-1), a proinflammatory chemokine important in monocyte recruitment into the vascular wall, was higher in lesions from apoE-/-VDR-/- mice, while αSMA showed reduced immunoreactivity in the fibrous cap of DKO animals (S3A, S3B, S3C, S3D, S3G, S3H, S3I and S3J Fig). To assess the role of VDR in plaque’s apoptosis, aortic arch tissue cross-sections were examined using the terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL) method. We did not detect a difference in the number of TUNEL-positive cells in atheromas from apoE-/-VDR-/- mice compared with apoE-/- littermates (S3E and S3F Fig).


Impaired Vitamin D Signaling in Endothelial Cell Leads to an Enhanced Leukocyte-Endothelium Interplay: Implications for Atherosclerosis Development.

Bozic M, Álvarez Á, de Pablo C, Sanchez-Niño MD, Ortiz A, Dolcet X, Encinas M, Fernandez E, Valdivielso JM - PLoS ONE (2015)

VDR ablation affects atheroma composition in apoE-/- mice.Tissue sections of aortic arch (A, B) and aortic root (D, E) from apoE-/- and apoE-/-VDR-/- mice were stained with anti-Mac-3 to quantify plaque macrophage content. (C) Staining intensity and % of Mac-3 positive cells in the aortic arch were graded semiquantitatively and histological scores were obtained from every sample, as explained in Materials and Methods. (F) Quantification of Mac-3 staining in the aortic root is shown as percentage of stained area relative to the area occupied by atheroma. The photomicrographs (20x magnification) show representative images of aortic arch and aortic root sections. Differences between groups were evaluated by the Student t test. Data presented are mean ± SEM of 10 mice/group. *p<0.05, *#p<0.001 vs. apoE-/- mice.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4556440&req=5

pone.0136863.g005: VDR ablation affects atheroma composition in apoE-/- mice.Tissue sections of aortic arch (A, B) and aortic root (D, E) from apoE-/- and apoE-/-VDR-/- mice were stained with anti-Mac-3 to quantify plaque macrophage content. (C) Staining intensity and % of Mac-3 positive cells in the aortic arch were graded semiquantitatively and histological scores were obtained from every sample, as explained in Materials and Methods. (F) Quantification of Mac-3 staining in the aortic root is shown as percentage of stained area relative to the area occupied by atheroma. The photomicrographs (20x magnification) show representative images of aortic arch and aortic root sections. Differences between groups were evaluated by the Student t test. Data presented are mean ± SEM of 10 mice/group. *p<0.05, *#p<0.001 vs. apoE-/- mice.
Mentions: To determine how VDR affects the nature of plaques, we further investigated atherosclerotic plaque composition. The increased lesion areas of the aortic arch and aortic root of apoE-/-VDR-/- mice correlated with increased macrophage (Mac-3) infiltration in both aortic regions (Fig 5A–5F). Immunoreactivity for monocyte chemoattractant protein (MCP-1), a proinflammatory chemokine important in monocyte recruitment into the vascular wall, was higher in lesions from apoE-/-VDR-/- mice, while αSMA showed reduced immunoreactivity in the fibrous cap of DKO animals (S3A, S3B, S3C, S3D, S3G, S3H, S3I and S3J Fig). To assess the role of VDR in plaque’s apoptosis, aortic arch tissue cross-sections were examined using the terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL) method. We did not detect a difference in the number of TUNEL-positive cells in atheromas from apoE-/-VDR-/- mice compared with apoE-/- littermates (S3E and S3F Fig).

Bottom Line: Knockdown of VDR in endothelial cells (shVDR) led to endothelial cell activation, characterized by upregulation of VCAM-1, ICAM-1 and IL-6, decreased peripheral blood mononuclear cell (PBMC) rolling velocity and increased PBMC rolling flux and adhesion to the endothelium. shVDR cells showed decreased IκBα levels and accumulation of p65 in the nucleus compared to shRNA controls.In vivo, deletion of VDR led to significantly larger aortic arch and aortic root lesions in apoE-/- mice, with higher macrophage content. apoE-/-VDR-/-mice showed higher aortic expression of VCAM-1, ICAM-1 and IL-6 when compared to apoE-/-VDR+/+ mice.Our data demonstrate that lack of VDR signaling in endothelial cells leads to a state of endothelial activation with increased leukocyte-endothelial cell interactions that may contribute to the more severe plaque accumulation observed in apoE-/-VDR-/- mice.

View Article: PubMed Central - PubMed

Affiliation: Nephrology Research Department, IRB Lleida, Lleida, Spain.

ABSTRACT
Endothelial cell activation leading to leukocyte recruitment and adhesion plays an essential role in the initiation and progression of atherosclerosis. Vitamin D has cardioprotective actions, while its deficiency is a risk factor for the progression of cardiovascular damage. Our aim was to assess the role of basal levels of vitamin D receptor (VDR) on the early leukocyte recruitment and related endothelial cell-adhesion-molecule expression, as essential prerequisites for the onset of atherosclerosis. Knockdown of VDR in endothelial cells (shVDR) led to endothelial cell activation, characterized by upregulation of VCAM-1, ICAM-1 and IL-6, decreased peripheral blood mononuclear cell (PBMC) rolling velocity and increased PBMC rolling flux and adhesion to the endothelium. shVDR cells showed decreased IκBα levels and accumulation of p65 in the nucleus compared to shRNA controls. Inhibition of NF-κB activation with super-repressor IκBα blunted all signs of endothelial cell activation caused by downregulation of VDR in endothelial cells. In vivo, deletion of VDR led to significantly larger aortic arch and aortic root lesions in apoE-/- mice, with higher macrophage content. apoE-/-VDR-/-mice showed higher aortic expression of VCAM-1, ICAM-1 and IL-6 when compared to apoE-/-VDR+/+ mice. Our data demonstrate that lack of VDR signaling in endothelial cells leads to a state of endothelial activation with increased leukocyte-endothelial cell interactions that may contribute to the more severe plaque accumulation observed in apoE-/-VDR-/- mice. The results reveal an important role for basal levels of endothelial VDR in limiting endothelial cell inflammation and atherosclerosis.

No MeSH data available.


Related in: MedlinePlus